With the increasing studies of microRNAs, more and more research reported that a specific miRNA could target multiple targets of a signaling pathway of related pathways. In some of these papers, to elucidate the specificity between miRNA and targets, point mutation in the seed sequence of miRNA were applied. I wonder if this is reasonable? As if the mutated miRNA mimics (for transient expression) were used, the mimics may have got new targets instead of these studied in the papers. And if the point mutation was applied while cloning (for stable cell line establishment), I doubt that this may lead to inhibiting the mature of the miRNA, and thus there would be no such a mutated miRNA at all in the stable cell lines. This confuses me somewhat. I wish those who are good at or interested in miRNA study give some discussions and suggestions.
Hi
Personally I do not think that mutating the seed sequence is a good strategy for studying miRNA function. In fact, if you mutate the seed sequence you are giving rise to a new miRNA with different functions. Pay attention that some miRNAs are just 1 base different.
The best way to inhibit a miRNA would be to use antagonistic RNA molecules (antagomirs) or a synthetic RNA sponge harboring miRNA targets.
Best. Good luck
Paco
Paco again summed it up nicely. Of course the best way is a knockout. That is most of the time a little bit too much. Inhibitors may do most of the time the trick. Sponges are a convoluted issue. Some work, some don't. I worked with a published sponge to find out that it does not work. So, I am a sponge-sceptic.
@Thomas.... I am also a spongeskeptic..... but at least are those kind of "text-book" solutions that are in the usual "menu".
@Paco:Sometimes the menu looks great but the cook is not doing his job so well...
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